Table 2.

EBNA-3C-interacting cellular partners and their biological consequences.

Interacting cellular proteins	Experimental method(s)	Functional deregulation	Ref.
RBP-Jκ (CBF-1)	Coimmunoprecipitation using both EBNA-3C-stable as well as EBV-transformed cells	Competes with EBNA-2 for the interaction with RBP-Jκ and subsequently downregulates EBNA-2-mediated transactivation of the EBV Cp promoter for EBNA expression. The interaction has been shown to be responsible for in vitro B-cell transformation and EBNA-3C-mediated growth maintenance of LCLs	[47,154]
p53	In vitro using GST-fused proteins and coimmunoprecipitation using both ectopically expressed as well as EBV-transformed cells	Blocks p53-dependent transcriptional activation and subsequent apoptotic induction	[98]
E2F1	In vitro using GST-fused proteins and coimmunoprecipitation using both ectopically expressed as well as EBV-transformed cells	Inhibits its DNA-binding ability and, as a result, blocks E2F1 transcriptional activity, as well as apoptotic induction, in response to DNA damage. In addition, EBNA-3C enhances E2F1 degradation by recruiting ubiquitin–proteasome machinery	[97]
Cyclin A	Yeast two-hybrid, in vitro using GST-fused proteins and coimmunoprecipitation using both ectopically expressed as well as EBV-transformed cells	Enhances kinase activity of cyclin A/CDK2 complex and rescues p27-mediated inhibition of cyclin A/CDK2 kinase activity by decreasing the molecular association between cyclin A and p27 in EBV-transformed cells	[95,96]
Cyclin E	In vitro using GST-fused protein incubated with radiolabeled EBNA-3C	No direct correlation has been depicted. It is speculated that EBNA-3C also regulates cyclin E/CDK2 activity during S phase	[95]
Cyclin D1	In vitro using GST-fused proteins and coimmunoprecipitation using both ectopically expressed as well as EBV-transformed cells	Stabilizes via blocking polyubiquitination levels, increases kinase activity of cyclin D1/CDK6 complex, thereby increasing the phosphorylation status of pRb tumor-suppressor protein, which, in turn, facilitates its ubiquitin–proteasome-dependent degradation. EBNA-3C interaction with cyclin D1 also results in nuclear accumulation of cyclin D1	[90,95]
Cyclin D2	Coimmunoprecipitation using ectopically expressed cells	No functional correlation has been established	[90]
Cyclin D3	Coimmunoprecipitation using ectopically expressed cells	No functional correlation has been established	[90]
MDM2	In vitro using GST-fused proteins and coimmunoprecipitation using both ectopically expressed as well as EBV-transformed cells	Enhances MDM2 stabilization through blocking its ubiquitin–proteasome-mediated proteolysis or recruiting deubiquitination activity. EBNA-3C also recruits MDM2 E3 ligase activity towards p53 for facilitating its degradation	[31]
pRb	In vitro using GST-fused proteins and coimmunoprecipitation using ectopically expressed cells	Is destabilized in an ubiquitin–proteasome-dependent pathway by recruiting SCFSkp2 E3 ligase activity	[87]
Chk2	In vitro using GST-fused proteins and coimmunoprecipitation using ectopically expressed cells	Releases G2/M cell cycle blockage induced by nocodazole. The interaction results in a predominant phosphorylation of residue and subsequently sequesters in the Cdc25c at S216 cytoplasm through interaction with 14-3-3	[113]
GADD34	Yeast two-hybrid and coimmunoprecipitation using ectopically expressed cells	Promotes eIF2-α and, at the same phosphorylation at S51 time, blocks XBP1 activation and ATF6 cleavage of unfolded protein response components	[155]
Spi-1/Spi-B	In vitro GST pulldown using radiolabeled EBNA-3C protein	Recruits Spi-1/Spi-B transcription factors to enhance EBNA-2-mediated LMP-1 promoter activation in a RBP-Jκ-independent manner	[39]
SCFSkp2 molecules	In vitro using GST-fused protein incubated with radiolabeled EBNA-3C proteins and coimmunoprecipitation using ectopically expressed cells	Recruits SCFSkp2 E3 ligase activity towards many tumor-suppressor proteins, including p27KIP1 and pRb to enhance their ubiquitin–proteasome-dependent proteolysis	[93]
c-Myc	In vitro using GST-fused proteins and coimmunoprecipitation using both ectopically expressed as well as EBV-transformed cells	Increases its stability as well as c-Myc-mediated transcriptional activation	[92]
Prothymosin-α	In vitro using GST-fused proteins and coimmunoprecipitation using both ectopically expressed as well as EBV-transformed cells	Through interaction with prothymosin-α, EBNA-3C recruits p300 HAT activity, as well as other basal transcription factors, to modulate gene transcription	[40,156]
CtBP	Coimmunoprecipitation using both ectopically expressed as well as EBV-transformed cells	With the interaction with CtBP, EBNA-3C aids in chromatin remodeling and epigenetic suppression of p16INK4A gene expression	[17,54]
NM23-H1	Yeast two-hybrid, in vitro GST pulldown and coimmunoprecipitation using both ectopically expressed as well as EBV-transformed cells	Enhances nuclear localization, modulates its transcriptional activity as well as cell migration ability. EBNA-3C induces metastasis in a nude mouse model in cooperation with Nm23-H1	[61,62,157]
Gemin3/DP103	Yeast two-hybrid, in vitro using GST-fused proteins and coimmunoprecipitation using ectopically expressed as well as EBV-transformed cells	Stabilizes Gemin3 and promotes a complex formation between Gemin3 and p53, which, in turn, blocks p53-mediated transcriptional activation as well as apoptosis	[134,137]
p300	In vitro using GST-fused proteins incubated with radiolabeled EBNA-3C proteins	Regulates p300- and prothymosin-α-mediated acetylation of histone molecules in the nucleosomes and results in transcriptional activation. However, a precise functional consequence has not been demonstrated	[40,156]
HDAC1	In vitro using GST-fused proteins and coimmunoprecipitation using ectopically expressed cells	HDAC1 was shown to play a crucial role in EBNA-3C-mediated transcription repression. However, a direct biological phenomenon has not yet been described	[41,42]
HDAC2	GST pulldown and coimmunoprecipiation experiments using stable cells as well as LCLs	It has been suggested that prothymosin-α plays an important role in regulating the association between EBNA-3C and HDAC molecules, as well as corepressor complexes including mSinA and NCoR	[42]
mSin3A	GST pulldown and coimmunoprecipiation experiments using stable cells as well as LCLs	Forms a transcriptional repressor complex. However, a direct functional correlation has not been described yet	[42]
NCoR	GST pulldown and coimmunoprecipiation experiments using stable cells as well as LCLs	Like mSin3A, NCoR presents in the same transcriptional repressor complex and a direct functional correlation has not been found	[42]
ING4	In vitro using GST-fused proteins and coimmunoprecipitation using both ectopically expressed as well as EBV-transformed cells	EBNA-3C blocks its interaction with p53 and thereby inhibits p53-mediated transcriptional as well as apoptotic activities	[99]
ING5	In vitro using GST-fused proteins and coimmunoprecipitation using both ectopically expressed as well as EBV-transformed cells	Similar to ING4, EBNA-3C inhibits its interaction with p53, and as a result, blocks p53-mediated transcriptional as well as apoptotic activation	[99]
GSK-3β	Coimmunoprecipitation using ectopically expressed cells	Inhibits phosphorylation of cyclin D1 at T286 and subsequently accelerates nuclear accumulation of cyclin D1, which contributes to its increased stability	[90]
SUMO-1/3	Yeast two-hybrid, GST pulldown and coimmunoprecipitation using ectopically expressed cells	Recruits SUMO-1 and -3 activities for LMP-1 promoter activation coupled with EBNA-2	[37,158]
MRS18–2	Yeast two-hybrid, GST pulldown and coimmunoprecipitation using LCLs	Interacts with and promotes nuclear localization, resulting in the disruption of pRb–E2F1 complexes	[88]

GST: Glutathione S-transferase; HAT: Histone acetyltransferase; HDAC: Histone deacetyltransferase; LCL: Lymphoblastoid cell line.